We are dedicated to creating more potent therapeutic monoclonal antibodies in order to increase the efficacy of cancer treatment.
Our initial focus on proven therapeutic targets is mAbs to the PD-1/PD-L system which acts to release the brakes on the immune system’s response to tumors. This has been the most innovative and successful new anti-cancer therapy introduced in the last decade. Analysts predict revenues from this therapeutic target to peak at $35bn/year by 2025. A license between major Pharma companies for a late stage product has been completed for an upfront fee of $890M indicating the value of this target.
Our HD-mAbTM will improve on the current therapeutic efficacy of the system starting with a mAb with comparable therapeutic efficacy of the current approved products.
Homologous Dimerization Technology (HDTM)
The general principle for engineering an HD antibody is that a simple peptide can be added to a recombinant, humanized antibody in a fashion that does not alter antigen binding or target localization while imparting a secondary property of homologous dimerization. The peptide has an affinity for self-association at least two orders of magnitude less than that of the antibody’s affinity for its target antigen.
The peptide imparting homologous dimerization properties to antibodies was derived from an experiment of nature; in response to recurrent bacterial infection, mice made a clonal response that had higher binding, opsonization and phagocytosis ability and a much higher neutralization titer. In research aimed at elucidating the reason for higher therapeutic efficacy, the antibody was found to express homologous dimerization ability. This property was localized to a novel 26 mer sequence, the parent to the sequence we utilize in our recombinant antibody form.
Shown in the Figure 2 is the T15 HD sequence. The original HD-T15 was shown to contain a sequence of 20-24 amino acids, since reproduced in a synthetic peptide, that was responsible for HD activity. The peptide has unique conformational properties (see above) and had the ability to bind to itself (lower right panel).
HD peptide have been initially incorporated into antibodies by a site-specific chemical conjugation method, resulting in 1 peptide/Fab. We then evolved a method of peptide incorporation into an human IgG1 backbone allowing for CDR replacement.
As shown in Figure 3, HD-mAbs are monomeric in solution, only demonstrating dimerization ability when put into non-physiologic conditions (such as PEG). When bound to target antigen, the HD-peptide is exposed and is able to interact with a second HD-peptide on another antibody without the antibody first binding to a target antigen; this allows for many more antibodies to be incorporated into a lattice like formation then dictated by the number of antigenic determinants. Most importantly, the lattice formation results in cross-linking of antigen which can trigger many biological activities.
Since this initial discovery, higher therapeutic potency has been extended to multiple antigen-antibody systems (15 to date) with many, different potential therapeutic indications. Two of the most studied systems are targeting Her-2 and CD-20.
Therapeutic Properties of HD-Antibodies
A summary of key therapeutic properties enhanced by HD technology are summarized separately below using as an example Herceptin and Rituxin.
A key reason for the platform’s ability to increase potency is shown in the Figure 4. The antibody is a Herceptin Biosimilar. Shown are results from Biacore measurement of antibody affinity. The HD version of Herceptin has greatly increased binding compared to Herceptin while at the same time has a reduced off-rate, meaning it stays bound to the target longer. Most striking is the binding of the HD form of Herceptin is non-saturable, accumulating more and more antibody on the target over time. Antigen-specific binding is the most important property of a therapeutic antibody and is the sole property responsible for potency for many therapeutic antibodies such as Avastin (anti-VEGF).
The most significant limitation of antibody therapy is antigenic heterogeneity; low antigen expression of a targeted cancer is a hallmark of most antigen-antibody systems; indeed even after antibody therapy of high expressing tumors, antibody therapy can result in selection of resistant, low antigen expressing variants. As an example, Herceptin therapy of breast cancer is limited to the 25% of high expressing tumors. Shown in Figure 5 are human lymphoma cells with binding of Rituxin detected by flow cytometry; Rituxin identifies 2 primary populations of cancer cells, one with a low antigen density and a second with higher density; the latter would be susceptible to therapy, the former not. HD-Rituxin in contrast binds both populations well, enabling treatment of even low antigen expressing cancer cells.
Enhanced binding or avidity (or decreased off rate) can likely be achieved for most antigen-antibody systems. This alone is a beneficial characteristic for certain applications such as antibody based imaging and radio-immunotherapy where retention of antibody and its label at the tumor site should enhance tumor imaging and increase the radiation delivered. Certain other applications such as antibody-dependent cellular cytotoxicity (ADCC) and complement-mediated cytotoxicity (C’MC) may also be enhanced with HD Technology due to increased amounts of antibody deposited on cells. An easily extrapolated application is the potential enhancement of antibody mediated phagocytosis and opsonization. Since the original discovery of native HD antibody was within a model system of an acute bacterial infection where protection was afforded by increased opsonization and phagocytosis, we expect that engineered HD antibodies will also express this enhanced property. Thus, clearance of blood-borne and tissue occurring pathogens may be significantly enhanced by this approach. Receptor blockade and the opposite, receptor mediated signaling, with appropriate antibodies, may be also significantly enhanced by the increased persistence of the antibody on the target and cross-linking of receptors.
Triggering Apoptosis (Cell Suicide)
As a result of lattice formation, antigen targets can be cross-linked on the cell surface. Such cross-linking can trigger a variety of biological activities. A potential therapeutic property for antibodies is apoptosis; this property is rarely encountered in mAbs. As shown in Figure 6, Herceptin does not trigger substantial apoptosis (upper right quadrant) but leaves most cells viable (lower left quadrant) after 24 hours of incubation. In contrast, HD-Herceptin (upper panel) causes apoptosis of most cancer cells. Thus, a widely used therapeutic antibody has acquired a new therapeutic mechanism.
In Vivo Efficacy
As shown in Figure 7, we evaluated HD-antibodies for efficacy in a relevant human tumor model, initiating therapy in a nude mouse model of a low antigen expressing breast ca. (MCF-7), 7 days after injection of tumor. As shown, Herceptin showed no therapeutic effect, with tumor measurements the same as the control (blue line). In contrast, HD-Herceptin suppressed tumor growth dramatically (red line). The lack of activity of Herceptin in this model was expected as it does not recognize low antigen expressing breast cancer. Upon histological examination of tumors treated with HD-Herceptin, few viable cancer cells were found while inflammatory cells were in abundance. Key for human therapy extrapolation was the maximal active dose of HD-Herceptin was 10ug while Herceptin had no effect upto 100 ug/dose.
The most important way of creating differentiation in the marketplace is through increasing potency. How that is achieved can be distinctive from one antigen/antibody system to another; to date our experience indicates that lattice formation can lead to:
- Apoptosis (standalone therapeutic mAbs)
- Enhanced internalization (ADC applications)
- Receptor clearance (Antagonistic applications)
- Enhanced phagocytosis and clearance (bacterial and viral infections)
- Enhanced dwell time on target antigen (RIT applications)
Enhanced potency can be viewed from 2 perspectives: the ability to give less of the drug to achieve the same therapeutic effect with a better COGS or to achieve better therapeutic results with the same dose used in the first generation antibody – Either use creates a market dominating antibody.